Work in this laboratory is aimed at elucidating the mechanism of stimulus- response coupling mediated by Ca2+ and calmodulin. The calmodulin- regulated protein phosphatase, calcineurin, is used as a model system. It was previously shown that the catalytic subunit of calcineurin, calcineurin A, is composed of a "catalytic domain" and a regulatory domain itself composed of an inhibitory and calmodulinbinding domain. The "catalytic domain" was shown this year to contain at least three subdomains, the catalytic center, the calcineurin B-binding site and the interaction site(s) for the immunosuppressant/immunophilin complexes. The calcineurin B-binding site has been identified and its interaction with calcineurin B, synthesized in E. coli, is being studied by multidimensional NMR in collaboration with Drs. Ad Bax and Jacob Anglister (NIDDK). Another collaborative project with Dr. Schreiber (Harvard University) has been initiated to identify the interaction site(s) of calcineurin with immunosuppressants. These studies were undertaken to establish the molecular basis of the role of calmodulin and calcineurin in the Ca2+- mediated regulation of gene expression leading to T cell activation that was recently discovered by Dr. Schreiber and his colleagues. Another important development has resulted from a collaborative project with Drs. Bax and Ikura at NIH. This is the elucidation of the structure of the complex of calmodulin with a calmodulin-binding peptide. These studies helped to define structural requirements for calmodulin target protein interactions and revealed the importance of the flexibility of the central helix in this interaction.